Signals from the environment cross the plasma membrane along a chain of interacting proteins. Hundreds of cell membrane receptors for hormones, neurotransmitters, light and odorants are linked to intracellular enzymes and ion channels (effectors) through a family of GTP-binding proteins (G proteins) made up of alpha and betagamma subunits. The alpha subunit is known to activate some effectors. Work supported by this grant provided the first evidence that betagamma subunits could also regulate the activity of effectors. Thus, receptors coupled to G proteins can produce a bifurcating signal to initiate an interacting network of cellular responses. It is now clear that in addition to the core components needed to send a signal across the membrane (receptor, G protein, effector), other proteins bind to these components and modulate the response. Our long-term goal is to understand all the factors that determine how a signal is transmitted across the membrane and how it selectively regulates downstream cellular pathways. To achieve this goal, we must define the interacting surfaces of the known proteins and the mechanisms by which they control effector activity or assemble and localize signaling complexes. We must also complete the inventory of proteins that interact with and influence the signaling system. Such structural and mechanistic information is a necessary prerequisite for the design of reagents that will modulate cellular responses to external signals and will eventually correct abnormal signal transduction in diseased or malignant cells. The Specific Aims are: to test the hypothesis that betagamma activates different effectors through different combinations of binding sites on the outer circumference of the beta propeller and to use the information to develop inhibitors that block specific subsets of betagamma function; to define the sites on PLCbeta that bind betagamma and those that are essential for activation by betagamma; and to identify novel proteins that bind to the G protein signal transduction system and modulate its cellular function.